Use of divalent cations for inhibiting erythrocyte dehydration in vivo

Abstract

The present invention provides a chemical class of active agents to be used as efficacious drugs in preventing erythrocyte dehydration, thus helping ameliorate the symptoms of certain hemoglobinopathies. The active agents include a class of compounds that can deliver to an erythrocyte in vivo a divalent cation. Increased intracellular concentrations of the divalent cation in the erythrocyte block K—Cl cotransport preventing dehydration of the erythrocyte, thus improving erythrocyte survival. These agents are to be administered by any preferred route of administration including oral, intravenous and parenteral routes. These agents may be co-administered with any other anti-hemoglobinopathic compounds. The methodology is effective for both long term and short term therapy; may be employed prophylactically and/or therapeutically; and may be used in acute crisis clinical situations.

We claim:  
     
       1. A pharmaceutical composition, comprising: 
       a pharmaceutically acceptable agent that blocks K—Cl cotransport in an erythrocyte of a subject,  
       an anti-hemoglobinopathic agent other than said agent that blocks K—Cl cotransport in an erythrocyte of a subject, and  
       a pharmaceutically acceptable carrier.  
     
     
       2. A kit comprising: 
       a package housing a container containing a pharmaceutically acceptable agent that blocks in a subject K—Cl cotransport in erythrocytes of the subject, in an amount effective to inhibit erythrocyte cell dehydration, and  
       housing instructions for using said agent to treat erythrocyte cell dehydration, in a subject in need of such treatment.  
     
     
       3. A method for inhibiting erythrocyte cell dehydration in a subject, comprising: administering to a subject in need of such treatment a pharmaceutically acceptable agent that blocks in a subject K—Cl cotransport in erythrocytes of the subject, in an amount effective to inhibit erythrocyte cell dehydration in the subject. 
     
     
       4. The method of claim  3 , wherein the pharmaceutically acceptable agent is of the general formula:                    
       wherein X is a divalent cation, and  
       wherein R1 and R2 each is an aryl group or a heteroaryl group which can be unsubstituted, or substituted with substituents selected from the group consisting of (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl and (C 1 -C 6 ) alkynyl, —CN, —OR′, —SR′, —NO 2 , —NR′R′, halogen, amino acid, —R″, —C(O)R″, —C(S)R″, —C(O)OR″, —C(S)OR″, —C(O)SR″, —C(S)SR″, —C(O)N(R″) 2 , —C(O)C(O)R″, —C(S)C(O)R″, —C(O)C(S)R″, —C(S)C(S)R″, —C(O)C(O)OR″, —C(S)C(O)OR″, —C(O)C(S)OR″, —C(O)C(O)SR″, —C(S)C(S)OR″, —C(S)C(O)SR″, —C(O)C(S)SR″, —C(S)C(S)SR″, —C(O)C(O)N(R″) 2 , —C(S)C(O)N(R″) 2 , —C(O)C(S)N(R″) 2 , or —C(S)C(S)N(R″) 2 ,  
       wherein each R′ is independently selected from the group consisting of —H, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl and (C 1 -C 6 ) alkynyl, and  
       wherein each R″ is independently selected from the group consisting of —H, (C 1 -C 6 ) alkyl. (C 1 -C 6 ) alkenyl, (C 1 -C 6 ) alkynyl, (C 6 -C 20 ) aryl, (C 6 -C 20 ) substituted aryl, (C 6 -C 26 )alkaryl and substituted (C 6 -C 26 ) alkaryl, and the aryl and alkaryl substituents are each independently selected from the group consisting of —CN, —OR′, —SR′, —NO 2 , —NR′R′, halogen, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl, (C 1 -C 6 ) alkynyl and trihalomethyl.  
     
     
       5. The method of claim  4 , wherein at least one of R1 and R2 is 2-pyrrolidone. 
     
     
       6. The method of claim  5 , wherein the agent is:                    
     
     
       7. The method of claim  4 - 6 , wherein said divalent cation X is selected from the group consisting of Mg, Ca, Mn, Sr and Zn. 
     
     
       8. The method of claim  7 , wherein said divalent cation X is Mg. 
     
     
       9. The method of claim  3 - 6 , wherein said subject suffers from sickle cell disease. 
     
     
       10. The method of claim  3 - 6 , wherein said subject suffers from thalassemia. 
     
     
       11. The method of claim  3 - 6 , wherein said administration is per oral. 
     
     
       12. The method of claim  3 - 6 , wherein said administration is parenteral. 
     
     
       13. The method of claim  3 - 6 , wherein said administration is intravenous. 
     
     
       14. The method of claim  3 - 6 , further comprising administering an anti-hemoglobinopathic agent, other than said agent that blocks K—Cl cotransport in an erydhrocyte of a subject. 
     
     
       15. A method for inhibiting erythrocyte cell dehydration, comprising: 
       contacting an erythrocyte cell with an agent that blocks K—Cl cotransport in an erythrocyte, in an amount effective to inhibit erythrocyte cell dehydration,  
       wherein the agent is of the general formula:                    
       wherein X is a divalent cation, and  
       wherein R1 and R2 each is an aryl group or a heteroaryl group which can be unsubstituted, or substituted with substituents selected from the group consisting of (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl and (C 1 -C 6 ) alkynyl, —CN, —OR′, —SR′, —NO 2 , —NR′R′, halogen, amino acid, —R″, —C(O)R″, —C(S)R″, —C(O)OR″, —C(S)OR″, —C(O)SR″, —C(S)SR″, —C(O)N(R″) 2 , —C(O)C(O)R″, —C(S)C(O)R″, —C(O)C(S)R″, —C(S)C(S)R″, —C(O)C(O)OR″, —C(S)C(O)OR″, —C(O)C(S)OR″, —C(O)C(O)SR″, —C(S)C(S)OR″, —C(S)C(O)SR″, —C(O)C(S)SR″, —C(S)C(S)SR″, —C(O)C(O)N(R″) 2 , —C(S)C(O)N(R′) 2 , —C(O)C(S)N(R″) 2 , or —C(S)C(S)N(R″) 2 ,  
       wherein each R′ is independently selected from the group consisting of —H, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl and (C 1 -C 6 ) alkynyl, and  
       wherein each R″ is independently selected from the group consisting of —H, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl, (C 1 -C 6 ) alkynyl, (C 6 -C 20 ) aryl, (C 6 -C 20 ) substituted aryl, (C 6 -C 26 ) alkaryl and substituted (C 6 -C 26 ) alkaryl, and the aryl and alkaryl substituents are each independently selected from the group consisting of —CN, —OR′, —SR′, —NO 2 , —NR′R′, halogen, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl, (C 1 -C 6 ) alkynyl and trihalomethyl.  
     
     
       16. The method of claim  14 , wherein said anti-hemoglobinopathic agent inhibits erythrocyte cell dehydration. 
     
     
       17. The method of claim  14 , wherein said anti-hemoglobinopathic agent inhibits sickle hemoglobin polymerization. 
     
     
       18. The method of claim  14 , wherein said anti-hemoglobinopathic agent increases fetal hemoglobin production. 
     
     
       19. The method of claim  14 , wherein said anti-hemoglobinopathic agent increases microvascular blood flow. 
     
     
       20. The method of claim  7 , further comprising administering an anti-hemoglobinopathic agent, other than said agent that blocks K-Cl cotransport in an erythrocyte of a subject. 
     
     
       21. The method of claim  20 , wherein said anti-hemoglobinopathic agent inhibits erythrocyte cell dehydration. 
     
     
       22. The method of claim  20 , wherein said anti-hemoglobinopathic agent inhibits sickle hemoglobin polymerization. 
     
     
       23. The method of claim  20 , wherein said anti-hemoglobinopathic agent increases fetal hemoglobin production. 
     
     
       24. The method of claim  20 , wherein said anti-hemoglobinopathic agent increases microvascular blood flow. 
     
     
       25. The method of claim  8 , further comprising administering an anti-hemoglobinopathic agent, other than said agent that blocks K-Cl cotransport in an erythrocyte of a subject. 
     
     
       26. The method of claim  8 , wherein said anti-hemoglobinopathic agent inhibits erythrocyte cell dehydration. 
     
     
       27. The method of claim  8 , wherein said anti-hemoglobinopathic agent inhibits sickle hemoglobin polymerization. 
     
     
       28. The method of claim  8 , wherein said anti-hemoglobinopathic agent increases fetal hemoglobin production. 
     
     
       29. The method of claim  8 , wherein said anti-hemoglobinopathic agent increases microvascular blood flow. 
     
     
       30. The method of claim  15 , wherein at least one of R1 and R2 is 2-pyrrolidone. 
     
     
       31. The method of claim  30 , wherein the agent is:                    
     
     
       32. The method of claim  15 , wherein said divalent cation X is selected from the group consisting of Mg, Ca, Mn, Sr and Zn. 
     
     
       33. The method of claim  32 , wherein said divalent cation X is Mg. 
     
     
       34. The method of claim  15 , further comprising contacting the erythrocyte cell with an anti-hemoglobinopathic agent, other than said agent that block K-Cl contransport in an erythrocyte. 
     
     
       35. The pharmaceutical composition of claim  1 , wherein the pharmaceutically acceptable agent that blocks K-Cl contransport in an erythrocyte is of the general formula:                    
       wherein X is a divalent cation, and  
       wherein R1 and R2 each is an aryl group or a heteroaryl group which can be unsubstituted, or substituted with substituents selected from the group consisting of (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl and (C 1 -C 6 ) alkynyl, —CN, —OR′, —SR′, —NO 2 , —NR′R′, halogen, amino acid, —R″, —C(O)R″, —C(S)R″, —C(O)OR″, —C(S)OR″, —C(O)SR″, —C(S)SR″, —C(O)N(R″) 2 , —C(O)C(O)R″, —C(S)C(O)R″, —C(O)C(S)R″, —C(S)C(S)R″, —C(O)C(O)OR″, —C(S)C(O)OR″, —C(O)C(S)OR″, —C(O)C(O)SR″, —C(S)C(S)OR″, —C(S)C(O)SR″, —C(O)C(S)SR″, —C(S)C(S)SR″, —C(O)C(O)N(R″) 2 , —C(S)C(O)N(R″) 2 , —C(O)C(S)N(R″) 2 , or —C(S)C(S)N(R″) 2 ,  
       wherein each R′ is independently selected from the group consisting of —H, (C 1 -C 6 ) alkyl,(C 1 -C 6 ) alkenyl and (C 1 -C 6 ) alkynyl, and  
       wherein each R″ is independently selected from the group consisting of —H, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl, (C 1 -C 6 ) alkynyl, (C 6 -C 20 ) aryl, (C 6 -C 20 ) substituted aryl, (C 6 -C 26 ) alkaryl and substituted (C 6 -C 26 ) alkaryl, and the aryl and alkaryl substituents are each independently selected from the group consisting of —CN, —OR′, —SR′, —NO 2 , —NR′R′, halogen, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl, (C 1 -C 6 ) alkynyl and trihalomethyl.  
     
     
       36. The pharmaceutical composotion of claim  35 , wherein at least one of R1 and R2 is 2-pyrrolidone. 
     
     
       37. The pharmaceutical composition of claim  36 , wherein the agent is:                    
     
     
       38. The pharmaceutical composition of claim  35 , wherein said divalent cation X is selected from the group consisiting of Mg, Ca, Mn, Sr and Zn. 
     
     
       39. The pharmaceutical composition of claim  38 , wherein said divalent cation X is Mg. 
     
     
       40. The pharmaceutical composition of claim  1 , wherein said pharmaceutically acceptable carrier is a comestible product. 
     
     
       41. The pharmaceutical composition of claim  35 , wherein said pharmaceutically acceptable carrier is a comestible product. 
     
     
       42. The pharmaceutical composition of claim  39 , wherein said pharmaceutically acceptable carrier is a comestible product. 
     
     
       43. The pharmaceutical composition of claim  1 , wherein said anti-hemoglobinopathic agent is selected from the group consisting of an erythrocyte cell dehydration inhibitor, a sickle hemoglobin inhibitor, an agent that increases fetal hemoglobin production and an agent that increases microvascular blood flow. 
     
     
       44. The pharmaceutical composition of claims  39 , wherein said antihemoglobinopathic agent is selecfted from the group consisting of an erythrocyte cell dehydration inhibitor, a sickle hemoglobin inhibitor, an agent that increases fetal hemoglobin production and an agent that increases microvascular blood flow. 
     
     
       45. The kit of claim  2 , wherein said agent is of the general formula:                    
       wherein X is a divalent cation, and  
       wherein R 1  and R 2  each is an aryl group or a heteroaryl group which can be unsubstituted, or substituted with substituents selected from the group consisting of (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl and (C 1 -C 6 ) alkynyl, —CN, —OR′, —SR′, —NO 2 , —NR′R′, halogen, amino acid, -R″, —C(O)R″, —C(S)R″, —C(O)OR″, —C(S)OR″, —C(O)SR″, —C(S)SR″, —C(O)N(R″) 2 , —C(O)C(O)R″, —C(S)C(O)R″, —C(O)C(S)R″, —C(S)C(S)R″, —C(O)C(O)OR″, —C(S)C(O)OR″, —C(O)C(S)OR″, —C(O)C(O)SR″, —C(S)C(S)OR″, —C(S)C(O)SR″, —C(O)C(S)SR″, —C(S)C(S)SR″, —C(O)C(O)N(R″) 2 , —C(S)C(O)N(R″) 2 , —C(O)C(S)N(R″) 2 , or —C(S)C(S)N(R″) 2 ,  
       wherein each R′is independently selected from the group consisting of —H, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl and (C 1 -C 6 ) alkynyl, and  
       wherein each R″ is independently selected from the group consisting of —H, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl, (C 1 -C 6 ) alkynyl, (C 6 -C 20 ) aryl, (C 6 -C 20 ) substituted aryl, (C 6 -C 26 ) alkaryl and substituted (C 6 -C 26 ) alkaryl, and the aryl and alkaryl substituents are each independently selected from the group consisting of —CN, —OR′, —SR′, —NO 2 , —NR′R′, halogen, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkynyl and trihalomethyl.  
     
     
       46. The kit of claim  45 , wherein at least one of R1 and R2 is 2-pyrrolidone. 
     
     
       47. The kit of claim  46 , wherein the agent is:                    
     
     
       48. The kit of claim  45 , wherein said divalent cation X is selected from the group consisting of Mg, Ca, Mn, Sr, and Zn. 
     
     
       49. The kit of claim  48 , wherein said divalent cation X is Mg. 
     
     
       50. The kit of claim  2 , further comprising, a container containing an anti-hemoglobinopathic agent other than said agent that blocks K-C 1  cotransport in erythrocytes of the subject. 
     
     
       51. The kit of claim  45 , further comprising, a container containing an anti-hemoglobinopathic agent other than said agent that blocks K-C 1  cotransport in erythrocytes of the subject. 
     
     
       52. The kit of claim  49 , further comprising, a container containing an anti-hemoglobinopathic agent other than said agent that blocks K-C 1  cotransport n erythrocytes of the subject.